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DESI-Mass Spectrometry Imaging Investigation of Discrete

and Cassette Drug Dosed Tissues
Philippa J Hart,¹ Andreas Danhorn,² Mark Towers,¹ and Emmanuelle Claude¹
¹Waters Corporation, Wilmslow, UK, ²Imperial College, London, UK

Mass spectrometry imaging for the distribution of drugs

and their associated metabolites, a DMPK solution.

GOAL
Desorption electrospray ionization (DESI)
mass spectrometry imaging (MSI) was
used to analyse tissue sections from
cassette dosed drug metabolism and
pharmacokinetic (DMPK) models. The
ability to map the distribution of drugs and
their metabolites in the liver, alongside
endogenous lipid species is demonstrated.

Figure 1. DESI-MS images of the four drugs and endogenous lipid PC(34:2), [M+K]+ in liver
tissue sections from: 2 and 6 hour post-cassette dosed models, 2 and 6 hour discrete dosed
BACKGROUND models, and untreated control. Top left panel shows sample key.

Assessing the ability of drug candidates

to reach the desired target(s) in
preclinical animal models is an essential
step of the pharmaceutical discovery
and development process. Typically,
whole-body autoradiography (WBA) is
the method routinely used for assessing
pharmacokinetic and distribution properties
of a drug candidate.
This technique requires the introduction of
a radiolabel to the synthesis of the drug, and
then allows the visualization of the radiolabel
in a quantitative, sensitive manner. However,
as it relies on the detection of the radiolabel,
WBA is unable to distinguish between
the parent compound and any potential
metabolites that could also contain the
radiolabel. Furthermore, to synthesize the
radiolabeled compound often requires an alteration in the chemistry used,
and can be an expensive procedure.
Mass spectrometry imaging (MSI) permits the visualization of molecules in a
label-free and multiplexed approach. This provides the accurate localization
of drug candidates and potential metabolites simultaneously, without the
need for labeling. There is also potential for a better understanding of the
mechanism of action of drug candidates, and the ability to identify m/z
values for use as biomarkers and companion diagnostics. Multimodal MSI
approaches are known to give complementary information.¹ For example,
secondary ion mass spectrometry (SIMS) can be used to generate high
spatial resolution images (nanometer level) with low compound coverage,
and matrix assisted laser desorption ionization (MALDI) can be used for
better compound coverage with micrometer spatial resolution. Recently,
electrospray (ESI) type ionization techniques, such as desorption
electrospray ionization (DESI) and liquid extraction surface analysis
(LESA), have been implemented for MSI experiments.

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THE SOLUTION
In this experiment using an orally dosed mouse
model, one cohort of animals was cassette
dosed with four drugs (Terfenadine,
Olanzapine, Moxifloxacin, and Erlotinib, at 25,
10, 10 and 25 mg/kg respectively) and a second
cohort was only dosed with Olanzapine at
25 mg/kg. A further cohort was vehicle dosed
to act as a control. Animals from each cohort
were euthanized at either 2 or 6 hrs post
administration and liver sections were then taken.

DESI-MSI analysis was performed using

a Prosolia 2D DESI source with a Waters®
sprayer on a Waters Xevo® G2-XS QTof Mass
Spectrometer with data acquisition and
processing through Waters High Definition
Imaging (HDI®) v1.4 Software. Ion images
demonstrating the localization of the four drugs
as well as endogenous species (mainly lipid
species) were generated from a single imaging
experiment, as shown in Figure 1. The distribution
and signal intensity of ions across the sections
were consistent with those shown in previously
published work.² In all figures, where no ions Figure 2. DESI-MS images of Terfenadine and two of its metabolites
were detected, dotted outlines of the tissues (hydroxyterfenadine and carboxyterfenadine) in liver tissue sections from:
2 and 6 hour post-cassette dosed models, 2 and 6 hour discrete dosed
are shown for reference. All data were
models, and untreated control.
acquired using a step size of 150µm and
subsequently normalized.

Within the same experiment it was also

possible to detect some of the metabolites from
Olanzapine, Terfenadine, and Erlotinib. The ion
images for Terfenadine (m/z 472.32) and for
two of its metabolites: hydroxyterfenadine
(m/z 488.31) and carboxyterfenadine
(m/z 502.29) are displayed in Figure 2.

In addition to the information gained for the

distribution of the drugs themselves and their
metabolites, it is also possible to view the
distribution of molecular ions from endogenous
species. Figure 3 shows the varying distribution
of endogenous phosphatidylcholine lipid species
(as labeled on the individual images).

Figure 3. DESI-MS images of endogenous phosphatidylcholines in liver

tissue sections from: 2 and 6 hour post-cassette dosed models, 2 and 6
hour discrete dosed models and untreated control.

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Overlaying the drug, metabolite and lipid ion

images, Figure 4 shows RGB overlaid ion images,
allowing clear visualization of the different spatial
distribution of ions within different anatomical
microstructures in the liver. This has the potential
to help determine the biological and histological
impact of the drugs and metabolites throughout
the tissue, and provides an extra level of detail
not available with other techniques.

The specific localization of molecules and degree

of information obtained can be improved by the
acquisition of additional imaging datasets at
higher spatial resolution, which, in the case of
DESI-MSI is possible using either a consecutive
tissue section, or if using solvents of a low
abrasive nature, on the same tissue section, Figure 4. DESI-MS RGB overlay images of Erlotinib (red), PC (32:0), [M+K]+, endogenous
as shown in the 150 µm images pictured here. phosphatidylcholine (green), and Terfenadine (blue), in liver tissue sections from: 2 and 6 hour
post-cassette dosed models, 2 and 6 hour discrete dosed models, and an untreated control.

SUMMARY
■■ Multiple drugs of low molecular weight
References
1. Swales, J.G. et.al. Analytical Chemistry, 2014, 86, 8473-8480
were detected in liver tissue sections,
independently of any labeling methodology. 2. Swales, J.G. et.al. Scientific Reports, 2016, DOI: 10.1038/srep37648

This saves valuable time and cost.

■■ Simultaneous detection of drug molecular

Acknowledgements
We thank Dr Richard J.A. Goodwin and John G. Swales from the Drug Safety
ions, their metabolites and endogenous
& Metabolism group at AstraZeneca R&D for providing all the samples used
lipid species was obtained, all in a single in this technical brief.
acquisition. Again, this saves valuable time,
cost, and ultimately can provide a wealth of
information from a single section of tissue.

■■ Image overlay function, for ion images and

optical images, allows for precise correlation
of MS with anatomical features.

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